Cochlear gap junctions coassembled from Cx26 and 30 show faster intercellular Ca<sup>2+</sup> signaling than homomeric counterparts

Sun, Jianjun; Ahmad, Shoab; Chen, Shanping; Tang, Wenxue; Zhang, Yanping; Chen, Ping; Lin, Xiaobo

doi:10.1152/ajpcell.00341.2004

Cited by 130 publications

(174 citation statements)

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“…The propagation of Ca 2ϩ waves could have been mediated by cellular release of ATP and subsequent purinergic receptor activation (15). However, blockade of this pathway by combined application of suramin and ATP did not affect the properties of Ca 2ϩ waves initiated by IP 3 injections (Fig.…”

Section: Gjs In Supporting Cells Mediated the Propagation Of Ca 2؉ Wamentioning

confidence: 94%

“…The medium was changed the day after plating and every other day thereafter. Cochlear cryosections and organotypic cultures were processed for fluorescent immunolabeling according to our published protocol (15). Samples were mounted with a SlowFade antifading mounting kit (Electron Microscopy Sciences, Hatfield, PA).…”

Section: Immunolabeling Of Cxs In Cochlear Cryosections and Organotypicmentioning

confidence: 99%

“…The standard ratio imaging technique (16) was used to record intracellular ionic concentration changes. Details were given in our previous publications (15). The speed of Ca 2ϩ wave propagation was calculated as the time difference in the onset of the Ca 2ϩ waves divided by the distance that the wave traveled.…”

Section: Measurement Of Gj-mediated Intercellular Ionic Coupling By Two-mentioning

confidence: 99%

“…For characterizing dye diffusion pattern and IP 3 injection-induced Ca 2ϩ waves in organotypic cochlear cultures, we confirmed the presence of GJs after experiments by performing immunolabeling procedures as outlined above. The cell-impermeable fluorescent dye (PI) was injected into cells by the single-cell electroporation method based on the technique described by Haas et al (17), and details were given in our previous publication (15). D-myo-inositol 1,4,5-trisphosphate hexapotassium salt (IP 3 , 250 M) was dissolved in normal pipette internal solution (130 mM CsCl͞10 mM EGTA͞0.5 mM CaCl 2 ͞3 mM MgATP͞2 mM Na 2 ATP͞10 mM Hepes, pH 7.2).…”

Section: Assessment Of Gj-mediated Intercellular Biochemical Couplingmentioning

confidence: 99%

“…IP 3 was introduced intracellularly by forming the whole-cell recording configuration. Cells were normally bathed in Hanks' balanced salt solution supplemented with 20 M ATP and 200 M 8-(3-benzamido-4-methylbenzamido)-naphthalene-1,3,5-trisulfonic acid (suramin) to eliminate Ca 2ϩ waves mediated by purinergic receptors (15).…”

Section: Assessment Of Gj-mediated Intercellular Biochemical Couplingmentioning

confidence: 99%

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Gap junction-mediated intercellular biochemical coupling in cochlear supporting cells is required for normal cochlear functions

Zhang

Tang²,

Ahmad³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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134

136

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Dysfunction of gap junctions (GJs) caused by mutations in connexin26 (Cx26) and Cx30 accounts for nearly half of all cases of hereditary nonsyndromic deafness cases. Although it is widely held that GJs connecting supporting cells in the organ of Corti mainly provide ionic pathways for rapid removal of K ؉ around the base of hair cells, the function of GJs in the cochlea remains unknown. Here we show that GJs were not assembled in the supporting cells of the organ of Corti until 3 days after birth in mice and then gradually matured to connect supporting cells before the onset of hearing. In organotypic cochlear cultures that were confirmed to express GJs, GJs mediated the propagation of intracellular Ca 2؉ concentration waves in supporting cells by allowing intercellular diffusion of inositol 1,4,5-trisphosphate. We found that a subset of structurally mild Cx26 mutations located at the second transmembrane region (V84L, V95M, and A88S) and a Cx30 mutation located at the first cytoplasmic segment (T5M) specifically affect the intercellular exchange of larger molecules but leave the ionic permeability intact. Our results indicated that Cx26 and Cx30 mutations that are linked to sensorineural deafness retained ionic coupling but were deficient in biochemical permeability. Therefore, GJ-mediated intercellular exchange of biochemically important molecules is required for normal cochlear functions.connexin ͉ deafness ͉ mutation

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Section: Gjs In Supporting Cells Mediated the Propagation Of Ca 2؉ Wamentioning

confidence: 94%

Section: Immunolabeling Of Cxs In Cochlear Cryosections and Organotypicmentioning

confidence: 99%

Section: Measurement Of Gj-mediated Intercellular Ionic Coupling By Two-mentioning

confidence: 99%

Section: Assessment Of Gj-mediated Intercellular Biochemical Couplingmentioning

confidence: 99%

Section: Assessment Of Gj-mediated Intercellular Biochemical Couplingmentioning

confidence: 99%

See 3 more Smart Citations

Gap junction-mediated intercellular biochemical coupling in cochlear supporting cells is required for normal cochlear functions

Zhang

Tang²,

Ahmad³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

134

136

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Gap Junctions and Connexins: The Molecular Genetics of Deafness

Nickel

Forge

2010

Encyclopedia of Life Sciences

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Mutations in the gene GJB2 encoding the gap‐junction protein connexin 26 ( Cx26 ), in particular, and in GJB6 coding for connexin 30 ( Cx30 ) are the most common cause of autosomal recessive sensorineural hearing loss in many world populations. Variants of GJB2 are also associated with dominant forms of both nonsyndromic and syndromic deafness. A complex picture of the roles of gap junctions in cochlear physiology has emerged. Rather than being mere conduits for the circulation of potassium ions in the inner ear, gap junctions have been implicated in the maintenance of metabolic homeostasis and in intercellular signalling among nonsensory cells. Studies of mutant channels and mouse models for connexin ‐related deafness have provided valuable insights into the heterogeneous mechanisms by which connexin mutations may cause cochlear dysfunction. Despite recent advances it is still not fully understood what roles gap junctions play in the inner ear and how connexin mutations cause deafness. Key Concepts: GJB2 and GJB6 have been mapped to the DFNB1 locus, which accounts for up to 50% of all cases of autosomal recessive nonsyndromic hearing loss. Molecular genetic tests for DFNB1 should include DNA sequencing of the GJB2 exons and mutation analysis for GJB6 deletions. The identification of factors underlying the phenotypic variability of connexin‐related hearing loss may improve clinical diagnosis and genetic counselling. A better understanding of the role of gap‐junctional communication in the inner ear and the structure–function relationships of connexin proteins is required for the development of mechanism‐based treatments of connexin‐associated hearing loss.

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Clinical and biophysical characterization of 19 GJB1 mutations

Tsai

Yang

Liao

et al. 2016

Ann Clin Transl Neurol

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ObjectiveCharcot–Marie–Tooth disease type X1 (CMTX1), which is caused by mutations in the gap junction (GJ) protein beta‐1 gene (GJB1), is the second most common form of Charcot–Marie–Tooth disease (CMT). GJB1 encodes the GJ beta‐1 protein (GJB1), which forms GJs within the myelin sheaths of peripheral nerves. The process by which GJB1 mutants cause neuropathy has not been fully elucidated. This study evaluated the biophysical characteristics of GJB1 mutants and their correlations with the clinical features of CMTX1 patients.MethodsAll patients with a validated GJB1 mutation were assessed using the Charcot–Marie–Tooth disease neuropathy score version 2 (CMTNS). The impacts of the mutations on the biophysical functions of GJB1 were characterized by assessing intracellular localization, expression patterns, and GJ Ca2+ permeability.ResultNineteen GJB1 mutations were identified in 24 patients with a clinical diagnosis of CMT. Six are novel mutations: p.L6S, p.I20F, p.I101Rfs*8, p.F153L, p.R215P, and p.D278V. Diverse pathological effects of the mutations were demonstrated, including reduced GJB1 expression, intracellular mislocalization, and altered GJ functions. GJB1 mutations that caused a complete loss of GJ Ca2+ permeability appeared to be associated with an earlier disease onset, whereas those resulting in preservation of GJ permeability and with predominant cell membrane expression tended to have a later onset and a milder phenotype.InterpretationThis study demonstrated that the degree of loss of GJ function caused by the GJB1 mutations might contribute to the onset and severity of neuropathic symptoms in CMTX1.

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Cochlear gap junctions coassembled from Cx26 and 30 show faster intercellular Ca²⁺ signaling than homomeric counterparts

Abstract: . Cochlear gap junctions coassembled from Cx26 and 30 show faster intercellular Ca 2ϩ signaling than homomeric counterparts.

Cited by 130 publications

References 42 publications

Gap junction-mediated intercellular biochemical coupling in cochlear supporting cells is required for normal cochlear functions

Gap junction-mediated intercellular biochemical coupling in cochlear supporting cells is required for normal cochlear functions

Gap Junctions and Connexins: The Molecular Genetics of Deafness

Clinical and biophysical characterization of 19 GJB1 mutations

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Cochlear gap junctions coassembled from Cx26 and 30 show faster intercellular Ca2+ signaling than homomeric counterparts

Abstract: . Cochlear gap junctions coassembled from Cx26 and 30 show faster intercellular Ca 2ϩ signaling than homomeric counterparts.

Cited by 130 publications

References 42 publications

Gap junction-mediated intercellular biochemical coupling in cochlear supporting cells is required for normal cochlear functions

Gap junction-mediated intercellular biochemical coupling in cochlear supporting cells is required for normal cochlear functions

Gap Junctions and Connexins: The Molecular Genetics of Deafness

Clinical and biophysical characterization of 19 GJB1 mutations

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Product

Resources

About

Cochlear gap junctions coassembled from Cx26 and 30 show faster intercellular Ca²⁺ signaling than homomeric counterparts